1. Field of the Invention
This invention relates to a method of reducing pyrogenicity of immobilized matrices used in the production and/or purification of biological and pharmaceutical products. More particularly, the invention relates to a method of inactivating pyrogens which contaminate immobilized matrices used in the production and/or purification of biological and pharmaceutical products by contacting the matrices in a packed bed configuration with a pyrogen inactivating solution.
2. Description of the Prior Art
Pyrogens are lipopolysaccharides, also known as endotoxins, which are an integral component of the outer cell surface of gram-negative bacteria. The pyrogens are shed from living bacteria and also are released into the environment when bacteria die and decompose. Since gram-negative bacteria are found universally in air, water and soil, bacterial endotoxins commonly contaminate the raw materials used to produce biological and pharmaceutical products as well as the processing systems and equipment used in the production of such products.
Pyrogens have numerous biological activities which include the production of fever, activation of clotting mechanisms and induction of shock. Consequently, it is essential that causative bacteria be inactivated in, and pyrogenic substances removed from, biological and pharmaceutical products as well as from the processing systems and equipment used in the production of the products.
Inactivation of pyrogens in products, processing systems and equipment, in general, includes: acid and base hydrolyses coupled with heat treatment; oxidation by hydrogen peroxide; alkylation by acetic anhydride, succinic anhydride and ethylene oxide; treatment with dry and moist heat and ionizing radiation. Removal of pyrogens include: rinsing equipment or materials with water; distillation; ultrafiltration; adsorption to activated carbon and removal by affinity chromatography. Methods of inactivation and removal of pyrogens must be compatible with the particular product and/or equipment and should not adversely affect the same while eliminating the pyrogenic substances. For example, while dry heat treatment destroys pyrogens, it is not suitable for heat-sensitive items, such as most polymerics, aqueous solutions and thermolabile biological agents. Wet heat, on the other hand, up to autoclave temperatures, does not destroy pyrogens, although it kills the organisms responsible for their production. Depyrogenation by the use of certain chemicals is also undesirable. For example, pyrogens are destroyed by sodium hydroxide at pH values greater than 8.0, but such high pH's are incompatible with various pharmaceutical processes as well as some products. Specifically: silicon based chromatography matrices dissolve; magnesium surfaces are violently attacked; and sensitive biological molecules tend to be unstable under these conditions.
The present invention is directed to the depyrogenation of solid matrices that are used in the processing of certain pharmaceutical/biological materials in order to prevent pyrogen-producing organisms and pyrogens produced thereby from entering into the process stream and products. Specific prior art methods and procedures do not relate to such depyrogenation and the general methods for depyrogenating products, equipments and raw materials are less than satisfactory to achieve Applicants' objectives.